The phylum Nematoda encompasses a group of unsegmented worms, of which approximately 2000 are known plant parasites. Nematodes parasitize the roots of a diverse collection of important food plants, including tomato, bean, sugar cane, peach, banana, pineapple and citrus.
In a well-balanced ecosystem, the multiplication of nematodes is checked by a variety of natural phenomena, including organisms which parasitize the nematode. In a cultivated field, however, nematodes may proliferate excessively, to the point of harming or destroying the crop.
The identification of bacterial parasites which infest nematodes suggested that these bacteria could be exploited for the control of nematodes in agriculture. One example of such a microbial nematicide is Pasteuria penetrans (also known as Bacillus penetrans), which is a bacterial endoparasite of several economically important namatodes, including Pratylenchus scribneri and four species of Meloidogyne. (Mankau, R. et al, 1977 J. Nematol. 9, 40-45.)
P.penetrans controls the multiplication of nematodes principally by destroying their reproduction. Spores of P. penetrans attach to the cuticle, or outer coat, of the developing female nematode, penetrate into its body cavity, and replicate therein. Infestation with multiplying P. penetrans prevents any significant nematode reproduction because physiological processes of the infected female nematode are taken over to promote P. penetrans reproduction. In a secondary form of biocontrol, spores infect a nematode, weakening it, and thereby reducing its infective capacity or rendering it susceptible to attack by other parasites and to pathogens (Davies, K. G. et al, 1988 Ann. Appl. Biol. 112, 491-501).
In order to exploit the bacterium P. penetrans as a biocontrol agent, a source for the bacteria is needed. Many types of bacteria can be grown under in vitro culture conditions (see for example, Bashan, Y. 1986 Appl. Env. Microbiol. 51, 1089-1098). However, since P. penetrans is an obligate parasite of nematodes, in vitro culture conditions have not been found to support the bacterial life cycle, and bacterial spores must be grown in and obtained from nematodes. Air-dried preparations of P. penetrans spores have been obtained from nematode-infested plant roots and tested for potential commercial use (Stirling, G. R., et al. 1980 Nematologica 26, 308-312; Stirling, G. R. 1984 Phytopathology 74, 55-60; Dube, B., et al, 1987 J. Nematol. 19, 222-227). Such preparations are limited in usefulness because the air-dried preparations are likely to contain viable nematodes or their eggs, which would further infest any field to which the spores were applied. Moreover, the powdery root preparations are not compatible with agricultural machinery. Clearly, alternative means for the agricultural application of microbial nematicides are needed.
Microencapsulation has been used for the packaging of living organisms within bead-like gels (Connick, W. J., Jr. 1988, In: Pesticide Formulations: Innovations and Development, Ed: B. Cross and H. B. Scher, pp. 241-250). For instance, sodium alginate solutions, upon exposure to metal cations such as calcium, form bead-like gels in which organisms may be entrapped. Upon exposure to the proper environment, the entrapped material is released at a rate controlled by the original formulation of the gel. Alginate beads are biodegradable and non-polluting.
Alginate gel has been used for the microencapsulation of beneficial bacteria and fungi grown in culture systems which exclude undesired plant pathogens. Biocontrol fungi were grown on agar plates or in fermentation vessels, and the wet fungal biomass was incorporated into alginate granules (Walker, H. L., et al, 1983, Weed Sci. 31, 333-338; Lewis, J. A., 1987, Phytopathology 75, 774-777; Fravel, D. R., et al, 1985 Phytopathology 75, 774-777). Biocontrol granules containing viable fungi were applied to soil in order to kill weeds or plant pathogenic microorganisma (Walker, H. L., Supra; Lewis, J. A., Supra). Beneficial rhizosphere bacteria were grown in culture systems free of plant pathogens and incorporated into alginate biocontrol granules, which were sowed concomitantly with seeds in order to promote plant growth (Bashan, Y. 1986, Supra.